A Novel Regulator of Cancer Cell Migration and Invasion

描述(由申请人提供)：细胞骨架和细胞迁移的调节是癌症侵袭和转移的一个重要方面。该项目的重点是一种新的蛋白质，Nischarin，它在细胞迁移和侵袭中具有抑制作用。整合素参与细胞过程，包括肌动蛋白细胞骨架的组织、细胞生长和存活、细胞运动和细胞侵袭。我们已经确定了一种新的蛋白质，它与a5细胞质尾巴特异地相互作用；我们将这种蛋白质命名为Nischarin。最近，我们借助rac1的效应域突变体，证明了Nischarin选择性地抑制RAC驱动的通过p21激活的激酶(PAK)发挥作用的信号级联反应。这些观察结果表明，Nischarin影响RAC和PAK介导的细胞骨架调节过程。与此一致，Nischarin与PAK结合，并抑制PAK的激酶活性。此外，我们最近的数据表明，Nischarin siRNA刺激片状脂体的形成和细胞迁移，并激活PAK激酶，表明Nischarin是细胞运动的重要调节因子。该提案的目的是进一步研究新描述的PAK和Nischarin之间的生化联系及其在肿瘤细胞迁移和侵袭中的作用。在Aim I中，我们将通过过量或低表达Nischarin来探讨Nischarin在癌细胞迁移和侵袭中的作用。我们还将研究PAK和Nischarin在伪足和伪足中的时空定位和调控。在AIM II中，我们将研究Nischarin对PAK下游效应因子的影响，包括LIMK和Cofilin，并将研究这如何影响癌细胞的侵袭。目的：我们将通过活细胞成像研究片脂动力学来评价Nischarin对片脂形成的影响；我们将可视化片脂突出、片脂回缩和膜褶皱的影响。评价Nischarin和PAK在调节肿瘤细胞迁移和侵袭中的作用，将拓宽目前对肿瘤细胞中PAK信号的理解，并可能为治疗干预提供新的途径。

DESCRIPTION (provided by applicant): Regulation of the cytoskeleton and cell migration is an important aspect of cancer invasion and metastasis. This project focuses on a novel protein, Nischarin that has an inhibitory role in cell migration and invasion. Integrins have been implicated in cellular processes including the organization of actin cytoskeleton, cell growth and survival, cell motility and cell invasion. We have identified a novel protein that specifically interacts with the a5 cytoplasmic tail; we have termed this protein 'Nischarin'. Recently, with the help of effector domain mutants of Rac1, we demonstrated that Nischarin selectively inhibits Rac-driven signaling cascades that act through p21 activated kinase (PAK). These observations suggest that Nischarin affects the process of cytoskeletal regulation mediated by Rac and PAK. Consistent with this, Nischarin binds to PAK, and inhibits the kinase activity of PAK. Furthermore, our recent data indicate that Nischarin siRNA stimulates lamellipodia formation and cell migration, and activates PAK kinase, suggesting that Nischarin is an important regulator of cell motility. The intent of the proposal is to investigate further the newly described biochemical connection between PAK and Nischarin and its role in tumor cell migration and invasion. In Aim I we will address the role of Nischarin in the migration and invasion of carcinoma cells by over or underexpressing Nischarin. Also we will examine spatio-temporal localization and regulation of PAK and Nischarin in pseudopodia and invadopodia. In Aim II, we will study the effect of Nischarin on downstream effectors of PAK including LIMK and cofilin, and will examine how this affects invasion of cancer cells. In Aim we will evaluate Nischarin's effects on lamellipodia formation by studying lamellipodial dynamics with live cell imaging; we will visualize the effects on lamellipodial protrusion, lamellipodial retraction and membrane ruffles. Evaluation of the role of Nischarin and PAK in the regulation of tumor cell migration and invasion will broaden the current understanding of PAK signaling in carcinoma cells, and may provide new avenues for therapeutic intervention.